Fig. 4: The prenatal Down syndrome brain exhibits altered differentiation trajectories and transposable element de-repression.

a Uniform Manifold Approximation and Projection (UMAP) visualization of RNA velocity trajectory analyses depicting developmental trajectories of cell types within mid-gestational euploid and Down syndrome (DS) brains from the single-nucleus (sn)RNA-seq dataset. Each arrow depicts the estimated direction and magnitude of change between cell states. b Latent time projection based on RNA velocity, capturing the relative progression of cells along pseudotemporal differentiation axes within euploid and DS brains. c Volcano plots of differential protein abundance from the proteomics analysis (n = 3 DS, n = 4 euploid). Each dot represents a protein and dots are colored according to enrichment: significantly upregulated (log₂FC ≥ 0.25, FDR ≤ 0.05) in teal, significantly downregulated (log₂FC ≤ 0.25, FDR ≤ 0.05) in purple, and non-significant genes in gray. Comparison is between DS and euploid samples. The horizontal line depicts FDR = 0.05, and the vertical lines depict log₂FC = 0.25. d Barplot of normalized enrichment score (NES) for select gene ontology (GO) biological processes from the proteomics analysis. Positive NES values indicate pathways enriched among upregulated proteins, while negative NES values correspond to pathways enriched among downregulated proteins in DS vs euploid. Bars are colored according to the adjusted p-value (FDR), reflecting the statistical significance of the enrichment. e Heatmap depicting the NES of transposable element (TE) subclasses in each cell subtype identified in the snRNA-seq dataset. Color reflects NES values, with teal indicating positive enrichment and purple indicating negative enrichment. Comparison is between DS and euploid samples. Gray represents enrichment scores not computed by GSEA. Asterisks (*) denote an FDR ≤ 0.05. f Immunostaining for LINE1-ORF1 (red) in SOX2⁺ (green) and DAPI-labeled nuclei (cyan) cells located near the ventricular zone (VZ) in prenatal human samples. Images are representative of those observed in mid-gestational prenatal euploid and DS brains. Scale bar is 30 μm. g Bar plots displaying LINE1-ORF1 intensity in SOX2⁺ cells, measured in arbitrary units (arb. units). Each dot represents the average intensity for each prenatal human brain (n = 3 DS, n = 3 euploid). Bars indicate the average intensity per condition, with error bars representing the standard error of the mean. Statistical significance was assessed using a two-sided t-test; p-value = 0.022.